1. Field of the Invention
The present invention relates to a vector and a method for detecting the action of a test substance on the gene transcription by activation of the ligand-dependent transcription factor corresponding to the test substance.
2. Description of the Related Art
Along with the progress in human domestic and industrial activities, there exists a serious social problem of adverse influences on the body by the chemical substances released into the environment. Currently, there are estimated to be at least 100,000 types of chemicals that are commercially produced, and in addition, there are still various types of chemicals that are unintendedly released into the environment in the human productive activities. The influence of most of these chemicals on the human body is unknown, and there is a concern about the diseases caused by these chemicals.
Among the chemicals present in the environment, the influence of the chemicals acting on nuclear receptors, i.e., ligand-dependent transcription factors, on the human body is more serious. The ligand-dependent transcription factors are proteins that have a function to change the transcription efficiency of a target gene on chromosome, wherein the proteins are activated by binding to a substrate, and bind to the recognition sequence present in the transcriptional regulatory region of the target gene. These ligand-dependent transcription factors play important roles in maintenance of homeostasis, reproduction, development, cell differentiation, and drug metabolism of organisms. Improper transcriptional regulation of a target gene by a transcription factor is known to cause abnormality in the expressed amount of the gene and hence various diseases. A method for measuring the activity of a chemical substance on the transcriptional regulatory potential of a ligand-dependent transcription factor is under development, for the purpose of restricting intake of chemical substances that induce diseases as a result of improper transcriptional regulation, or for the treatment and prevention of the diseases caused by such chemical substances, by detecting these chemical substances from among the chemicals present in the environment.
For example, for measurement of the activity of a test substance on the transcriptional regulatory potential, known is a method for measuring the activity of a test substance on the transcriptional regulation of a ligand-dependent transcription factor, by introducing a vector containing a reporter gene linked to downstream of the recognition sequence of a ligand-dependent transcription factor, and measuring the expressed amount of the reporter gene after the cell is incubated in the presence of the test substance. For measurement of the activity of a test substance similar to polycyclic aromatic hydrocarbons (PAHs) in structure, there has been proposed a method of measuring the activity of a test substance on the aryl hydrocarbon receptor (AhR)-mediated transcriptional regulation, by using a cell containing a vector having a reporter gene linked to downstream of a recognition sequence (i.e., xenobiotic responsive element (XRE)) for an aryl hydrocarbon receptor (AhR) which uses the PAHs as substrates. (JP-A 2000-253889 (KOKAI) and U.S. Pat. No. 5,854,010). Alternatively, for measurement of the influence of a test substance on a reproductive function, there has been proposed a method of measuring the activity of a test substance on female-hormone (estrogen) receptor (ER) or male-hormone (androgen) receptor (AR)-mediated transcription regulation, by using a cell containing a vector having a reporter gene linked to downstream of a recognition sequence for ER, i.e., estrogen responsive sequence (ERE), or a recognition sequence for AR, i.e., androgen responsive sequence (ARE) (JP-A 2002-253231 (KOKAI)). ER and AR are ligand-dependent transcription factors for controlling the reproductive function.
However, these methods only allow detection of the influence of the test substance on only one kind of ligand-dependent transcription factor-mediated transcriptional activation, by using one type of vector. In other words, these methods allow detection of the influence of the test substance on ligand-dependent transcription factor-mediated transcriptional activation, only by detecting separately the binding of each of activated transcription factors AhR, ER and AR to the corresponding recognition sequences XRE, ERE and ARE.